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Novel candidate genes influencing natural variation in potato tuber cold sweetening identified by comparative proteomics and association mapping.

Fischer M, Schreiber L, Colby T, Kuckenberg M, Tacke E, Hofferbert HR, Schmidt J, Gebhardt C - BMC Plant Biol. (2013)

Bottom Line: The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips.We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS.Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Plant Breeding and Genetics, Max-Planck-Institute for Plant Breeding Research, Cologne, Germany. fischer@mpipz.mpg.de

ABSTRACT

Background: Higher plants evolved various strategies to adapt to chilling conditions. Among other transcriptional and metabolic responses to cold temperatures plants accumulate a range of solutes including sugars. The accumulation of the reducing sugars glucose and fructose in mature potato tubers during exposure to cold temperatures is referred to as cold induced sweetening (CIS). The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips. CIS-tolerance varies considerably among potato cultivars. Our objective was to identify by an unbiased approach genes and cellular processes influencing natural variation of tuber sugar content before and during cold storage in potato cultivars used in breeding programs. We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS. DNA polymorphisms in genomic sequences encoding differentially expressed proteins were tested for association with tuber starch content, starch yield and processing quality.

Results: Pronounced natural variation of CIS was detected in tubers of a population of 40 tetraploid potato cultivars. Significant differences in protein expression were detected between CIS-tolerant and CIS-sensitive cultivars before the onset as well as during cold storage. Identifiable differential proteins corresponded to protease inhibitors, patatins, heat shock proteins, lipoxygenase, phospholipase A1 and leucine aminopeptidase (Lap). Association mapping based on single nucleotide polymorphisms supported a role of Lap in the natural variation of the quantitative traits tuber starch and sugar content.

Conclusions: The combination of comparative proteomics and association genetics led to the discovery of novel candidate genes for influencing the natural variation of quantitative traits in potato tubers. One such gene was a leucine aminopeptidase not considered so far to play a role in starch sugar interconversion. Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

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Protein spots showing differential expression between genotype pools CIS-t and CIS-s before and/or after cold treatment. A virtual tuber protein pattern was generated by fusing 2D-PAGE gel images from both genotype pools CIS-t and CIS-s at T0. Conditions for protein separation in the first and second dimension are shown on the right. The numbered arrow heads point to the position of the 50 differential proteins described in Table 2, Additional file 4: Table S4 and Additional file 5: Table S5. (A) Proteins from 40 to 200 kDa were separated on IPG strips with immobilized pH gradient of 3–11 in the first dimension (IEF) and by 10% Tris-glycine SDS-PAGE in the second dimension. (B) Proteins between 5 and 40 kDa were separated on pH 3–7 IPG strips in the first dimension and by 16% Tris-tricine polyacrylamide gels in the second dimension.
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Figure 3: Protein spots showing differential expression between genotype pools CIS-t and CIS-s before and/or after cold treatment. A virtual tuber protein pattern was generated by fusing 2D-PAGE gel images from both genotype pools CIS-t and CIS-s at T0. Conditions for protein separation in the first and second dimension are shown on the right. The numbered arrow heads point to the position of the 50 differential proteins described in Table 2, Additional file 4: Table S4 and Additional file 5: Table S5. (A) Proteins from 40 to 200 kDa were separated on IPG strips with immobilized pH gradient of 3–11 in the first dimension (IEF) and by 10% Tris-glycine SDS-PAGE in the second dimension. (B) Proteins between 5 and 40 kDa were separated on pH 3–7 IPG strips in the first dimension and by 16% Tris-tricine polyacrylamide gels in the second dimension.

Mentions: Based on the accumulation of reducing sugars during cold treatment (Figure 1B) 10 cultivars were selected, five each with the lowest (CIS-tolerant, CIS-t) and highest (CIS-sensitive, CIS-s) RSC values (indicated in Additional file 1: Table S1). Total soluble protein was extracted from tubers of the CIS-t and CIS-s cultivars prior to (T0) and after 2, 4 and 12 weeks of cold treatment (2w, 4w, 12w). At each time point two protein pools were created. The first pool was made of equimolar amounts of proteins from CIS-t cultivars and the second from CIS-s cultivars. The protein pools were separated by 2D-PAGE under two different conditions to resolve proteins of different size classes (Figure 3). By comparing protein profiles of the genotype pools CIS-t and CIS-s, protein spots were identified that showed quantitative as well as qualitative differences at one or more time points. Fifty protein spots (Figure 3) showed a minimum of a two-fold difference in spot intensity between genotype pools CIS-t and CIS-s at least at one time point (Table 2). The kinetics of the spot intensities during cold treatment and genomic positions of the corresponding genes are shown in Additional file 4: Table S4.


Novel candidate genes influencing natural variation in potato tuber cold sweetening identified by comparative proteomics and association mapping.

Fischer M, Schreiber L, Colby T, Kuckenberg M, Tacke E, Hofferbert HR, Schmidt J, Gebhardt C - BMC Plant Biol. (2013)

Protein spots showing differential expression between genotype pools CIS-t and CIS-s before and/or after cold treatment. A virtual tuber protein pattern was generated by fusing 2D-PAGE gel images from both genotype pools CIS-t and CIS-s at T0. Conditions for protein separation in the first and second dimension are shown on the right. The numbered arrow heads point to the position of the 50 differential proteins described in Table 2, Additional file 4: Table S4 and Additional file 5: Table S5. (A) Proteins from 40 to 200 kDa were separated on IPG strips with immobilized pH gradient of 3–11 in the first dimension (IEF) and by 10% Tris-glycine SDS-PAGE in the second dimension. (B) Proteins between 5 and 40 kDa were separated on pH 3–7 IPG strips in the first dimension and by 16% Tris-tricine polyacrylamide gels in the second dimension.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3750364&req=5

Figure 3: Protein spots showing differential expression between genotype pools CIS-t and CIS-s before and/or after cold treatment. A virtual tuber protein pattern was generated by fusing 2D-PAGE gel images from both genotype pools CIS-t and CIS-s at T0. Conditions for protein separation in the first and second dimension are shown on the right. The numbered arrow heads point to the position of the 50 differential proteins described in Table 2, Additional file 4: Table S4 and Additional file 5: Table S5. (A) Proteins from 40 to 200 kDa were separated on IPG strips with immobilized pH gradient of 3–11 in the first dimension (IEF) and by 10% Tris-glycine SDS-PAGE in the second dimension. (B) Proteins between 5 and 40 kDa were separated on pH 3–7 IPG strips in the first dimension and by 16% Tris-tricine polyacrylamide gels in the second dimension.
Mentions: Based on the accumulation of reducing sugars during cold treatment (Figure 1B) 10 cultivars were selected, five each with the lowest (CIS-tolerant, CIS-t) and highest (CIS-sensitive, CIS-s) RSC values (indicated in Additional file 1: Table S1). Total soluble protein was extracted from tubers of the CIS-t and CIS-s cultivars prior to (T0) and after 2, 4 and 12 weeks of cold treatment (2w, 4w, 12w). At each time point two protein pools were created. The first pool was made of equimolar amounts of proteins from CIS-t cultivars and the second from CIS-s cultivars. The protein pools were separated by 2D-PAGE under two different conditions to resolve proteins of different size classes (Figure 3). By comparing protein profiles of the genotype pools CIS-t and CIS-s, protein spots were identified that showed quantitative as well as qualitative differences at one or more time points. Fifty protein spots (Figure 3) showed a minimum of a two-fold difference in spot intensity between genotype pools CIS-t and CIS-s at least at one time point (Table 2). The kinetics of the spot intensities during cold treatment and genomic positions of the corresponding genes are shown in Additional file 4: Table S4.

Bottom Line: The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips.We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS.Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Plant Breeding and Genetics, Max-Planck-Institute for Plant Breeding Research, Cologne, Germany. fischer@mpipz.mpg.de

ABSTRACT

Background: Higher plants evolved various strategies to adapt to chilling conditions. Among other transcriptional and metabolic responses to cold temperatures plants accumulate a range of solutes including sugars. The accumulation of the reducing sugars glucose and fructose in mature potato tubers during exposure to cold temperatures is referred to as cold induced sweetening (CIS). The molecular basis of CIS in potato tubers is of interest not only in basic research on plant adaptation to environmental stress but also in applied research, since high amounts of reducing sugars affect negatively the quality of processed food products such as potato chips. CIS-tolerance varies considerably among potato cultivars. Our objective was to identify by an unbiased approach genes and cellular processes influencing natural variation of tuber sugar content before and during cold storage in potato cultivars used in breeding programs. We compared by two-dimensional polyacrylamide gel electrophoresis the tuber proteomes of cultivars highly diverse for CIS. DNA polymorphisms in genomic sequences encoding differentially expressed proteins were tested for association with tuber starch content, starch yield and processing quality.

Results: Pronounced natural variation of CIS was detected in tubers of a population of 40 tetraploid potato cultivars. Significant differences in protein expression were detected between CIS-tolerant and CIS-sensitive cultivars before the onset as well as during cold storage. Identifiable differential proteins corresponded to protease inhibitors, patatins, heat shock proteins, lipoxygenase, phospholipase A1 and leucine aminopeptidase (Lap). Association mapping based on single nucleotide polymorphisms supported a role of Lap in the natural variation of the quantitative traits tuber starch and sugar content.

Conclusions: The combination of comparative proteomics and association genetics led to the discovery of novel candidate genes for influencing the natural variation of quantitative traits in potato tubers. One such gene was a leucine aminopeptidase not considered so far to play a role in starch sugar interconversion. Novel SNP's diagnostic for increased tuber starch content, starch yield and chip quality were identified, which are useful for selecting improved potato processing cultivars.

Show MeSH
Related in: MedlinePlus